Faculty Bibliography

BACKGROUND:The evaluation of antibiotic immediate-type hypersensitivity is intricate because of nonstandardized skin testing and challenge method variability. OBJECTIVE:To determine the safety outcomes and risk factors for antibiotic challenge reactions in patients reporting a history of antibiotic immediate-type hypersensitivity. METHODS:A 5-year retrospective review of patients evaluated for immediate-type antibiotic allergy was conducted. Data analyzed included patient demographics, index reaction details, and outcomes of skin testing and challenges, classified as single-step or multistep. RESULTS:Antibiotic hypersensitivity history was identified in 211 patients: 78% to penicillins, 10% to fluoroquinolones, 7.6% to cephalosporins, and 3.8% to carbapenems. In total, 179 patients completed the challenges (median age 67 years, range 50-76 years, 56% women), and compared with nonchallenged patients, they reported nonanaphylactic (P < .001) and remote index (P = .003) reactions. Sixteen patients (8.9%) experienced challenge reactions (5 of 28 for single-step challenge, 11 of 151 for multistep challenge), and 11 of these patients had negative skin testing results before the challenge. Challenge-reactive patients were significantly younger (P = .007), more often women (P = .036), and had additional reported antibiotic allergies (P = .005). No correlation was detected between the reported index and observed challenge reaction severities (κ = -0.05, 95% confidence interval -0.34 to 0.24). Anaphylactic rates were similar during single-step and multistep challenges (3.6% vs 3.3%). CONCLUSION:In the present population, younger women with multiple reported antibiotic allergies were at greatest risk for challenge reactions. Negative skin testing results did not exclude reactions, and index severity was not predictive of challenge outcome. The multistep and full-dose methods demonstrated a comparable reaction risk for anaphylaxis.

Matrix metalloproteases MMP-2 and MMP-9 have been implicated in the physiologic catabolism of Alzheimer amyloid-beta (Abeta). Conversely, their association with vascular amyloid deposits, blood-brain barrier disruption, and hemorrhagic transformations after ischemic stroke also highlights their involvement in pathologic processes. To better understand this dichotomy, recombinant human (rh) MMP-2 and MMP-9 were incubated with Abeta40 and Abeta42 and the resulting proteolytic fragments assessed via immunoprecipitation and quantitative mass spectrometry. Both MMPs generated Abeta fragments truncated only at the C-terminus, ending at positions 34, 30 and 16. Using deuterated homologues as internal standards, we observed limited and relatively slow degradation of Abeta42 by rhMMP-2 while the enzyme cleaved >80% of Abeta40 during the first hour of incubation. rhMMP-9 was significantly less effective, particularly in degrading Abeta1-42, although the targeted peptide bonds were identical. Using Abeta1-34 and Abeta1-30, we demonstrated that these peptides are also substrates for both MMPs, cleaving Abeta1-34 to produce Abeta1-30 first and Abeta1-16 subsequently. Consistent with the kinetics observed with full-length Abeta, rhMMP-9 degraded only a minute fraction of Abeta1-34 and was even less effective in producing Abeta1-16. Further degradation of Abeta1-16 by either MMP-2 or MMP-9 was not observed even after prolonged incubation times. Notably, all MMP-generated C-terminally truncated Abeta fragments were highly soluble, did not exhibit fibrillogenic properties or induce cytotoxicity in human cerebral microvascular endothelial or neuronal cells supporting the notion that these truncated Abeta species are associated with clearance mechanisms rather than being key elements in the fibrillogenesis process.

Patients carrying mutations within the amyloid-beta (Abeta) sequence develop severe early-onset cerebral amyloid angiopathy with some of the related variants manifesting primarily with hemorrhagic phenotypes. Matrix metalloproteases (MMPs) are typically associated with blood brain barrier disruption and hemorrhagic transformations after ischemic stroke. However, their contribution to cerebral amyloid angiopathy-related hemorrhage remains unclear. Human brain endothelial cells challenged with Abeta synthetic homologues containing mutations known to be associated in vivo with hemorrhagic manifestations (AbetaE22Q and AbetaL34V) showed enhanced production and activation of MMP-2, evaluated via Multiplex MMP antibody arrays, gel zymography, and Western blot, which in turn proteolytically cleaved in situ the Abeta peptides. Immunoprecipitation followed by mass spectrometry analysis highlighted the generation of specific C-terminal proteolytic fragments, in particular the accumulation of Abeta-(1-16), a result validated in vitro with recombinant MMP-2 and quantitatively evaluated using deuterium-labeled internal standards. Silencing MMP-2 gene expression resulted in reduced Abeta degradation and enhanced apoptosis. Secretion and activation of MMP-2 as well as susceptibility of the Abeta peptides to MMP-2 degradation were dependent on the peptide conformation, with fibrillar elements of AbetaE22Q exhibiting negligible effects. Our results indicate that MMP-2 release and activation differentially degrades Abeta species, delaying their toxicity for endothelial cells. However, taking into consideration MMP ability to degrade basement membrane components, these protective effects might also undesirably compromise blood brain barrier integrity and precipitate a hemorrhagic phenotype